Failure of retinal ganglion cells (RGCs) to survive or regenerate their optic nerve axons underlies permanent visual disability in glaucoma, trauma, and other eye diseases. In this application we propose to study the role of a newly defined perinuclear cAMP second messenger compartment in neuroprotection and axon nerve regeneration. The phosphodiesterase PDE4D3 is specifically associated with the nuclear envelope-associated scaffold protein mAKAP?. Preliminary data show that cAMP signaling at mAKAP? in neurons is tightly regulated by PDE4D3 such that displacement of the PDE using a specific anchoring disruptor peptide promotes perinuclear protein kinase A activity and neurite extension in vitro and RGC survival in vivo following optic nerve crush, mimicking the application of exogenous cAMP analog. The mechanisms how cAMP at mAKAP? contributes to neuroprotection and neurite outgrowth are the focus of this project. For example, we propose that PDE4D3 serves as the fulcrum for crosstalk between cAMP and ERK5 pathways at mAKAP? signalosomes. In addition, potential effectors for mAKAP?-dependent cAMP-PKA neuroprotective signaling are class IIa HDACs (HDACs 4 and 5) that organize co-repressor complexes on chromatin via binding to transcription factors such as MEF2. PKA signaling promotes HDAC4/5 nuclear localization in other cell types, and we propose that mAKAP? and PKA-dependent HDAC phosphorylation results in HDAC nuclear localization in neurons promoting RGC survival following optic nerve injury. The central hypothesis of this project is as follows: Perinuclear cAMP signaling at PDE4D3-regulated mAKAP? signalosomes in neurons promotes class IIa HDAC nuclear localization, enhancing RGC survival and axon regeneration following optic nerve injury. Aim 1) Mechanisms underlying cAMP- dependent neuroprotective and axon regenerative signaling at mAKAP? signalosomes. Using a nuclear- envelope localized PKA activity reporter, we will determine by FRET imaging whether mAKAP?-bound PDE4D3 is regulated by neurotrophin-dependent ERK5 signaling and other upstream signals. The role of class IIa HDACs in neuron survival and axon growth will be studied using primary neuronal cultures, and whether HDAC intracellular localization is regulated by cAMP at mAKAP? signalosomes will be studied by live cell imaging. Aim 2) Targeting of perinuclear cAMP signaling as a therapeutic approach for RGC protection and optic nerve regeneration. To test whether intravitreal gene therapy targeting the mAKAP? compartment is synergistic with neurotrophin therapy that induces ERK5 signaling, adeno-associated virus vectors expressing a PDE4D3 anchoring disruptor peptide will be injected intravitreally before or after optic nerve crush surgery with or without simultaneous injection of BDNF. Histology and vision functional assays will be used to assess preservation and/or restoration of RGC/optic nerve function. To test whether enhanced class IIa HDAC nuclear localization confers neuroprotection, additional mice will be treated with adeno-associated virus vectors expressing mutant HDAC proteins and similarly studied for their role in optic nerve injury.